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Our Abpromise guarantee covers the use of ab32795 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use at an assay dependent dilution. PubMed: 19910498|
|IP||Use at an assay dependent dilution.|
|WB||Use a concentration of 2 µg/ml. Detects a band of approximately 92 kDa (predicted molecular weight: 86 kDa).|
|IHC-P||Use a concentration of 0.5 - 1 µg/ml.|
ab32795 staining TPX2 in HeLa cells and mouse NIH-3T3 cells (fuzzier pattern, different from the high-quality sharp signal seen in Human cells), by immunofluorescence.
optimal antibody dilution: 4µg/ml
optimal fixation protocol: PFA/Triton fixation: 10 min room at room temperature, in 3,7 % PFA diluted in PHEM buffer (45 mM Hepes pH 6,9, 45 mM Pipes pH 6,9, 5 mM MgCl2, 10 mM EGTA) containing 0.2% Triton X-100, followed by 3 washes in PBS - Alternative fixation protocol also gives good staining: 6 min in cold Methanol at -20°C, then 3 washes in PBS.
IF was performed following a standard protocol: Blocking, 30 min; primary antibody, 1 hr; secondary antibody, 45 min. All incubations were at 37 °C in PBS/ 0.1% Tween containing 3% BSA.
Image from Grover A et al.,PLoS One. 2012;7(1):e30890. Epub 2012 Jan 27. Fig 7.; doi:10.1371/journal.pone.0030890; January 27, 2012, PLoS ONE 7(1): e30890.